DNA repair Mechanisms

1. DNA Repair Mechanisms In Prokaryotes
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Shariqa Aisha
University of Kashmir
Department of Bioresources
Email id:
rasoolshariqa@gmail.com

2.  Introduction.
 DNA damage.
 Sources of DNA damage.
 DNA repair.
 Direct repair.
 Excision repair.
 Mismatch repair.

3.  DNA damage is an alteration in the chemical structure of DNA, such as a break in a
strand of DNA, a base missing from the backbone of DNA, or a chemically changed base
in a DNA.
 The vast majority of DNA damage affects the primary structure of the double helix.
Source of damage:
 Endogenous damage such as attack by reactive oxygen species produced from normal
metabolic byproducts ( spontaneous mutation), especially the process of oxidative
deamination.
 Depurination.
 Also includes replication errors.
 Exogenous damage caused by external agents such as_ ultraviolet radiation, x-rays,
gamma rays, viruses etc.

4.  DNA repair is a collection of process by which a cell identifies and corrects damage to
the DNA molecules that encode its genome.
 The DNA repair processes is constantly active as it responds to damage in the DNA
structure.
 Most cells posses three different categories of DNA repair systems:
i) Direct repair
i) Excision repair
ii) Mismatch repair

6.  Direct repair systems act directly on damaged
nucleotides, converting each one back to its original
structure.
 One common type of UV radiation mediated damages,
pyrimidine dimers, are repaired by a light-dependent
direct system called photoreactivation.
 In E. coli, the process involves the enzyme called DNA
photolyase.
 When stimulated by light with a wavelength between
300 and 500 nm, the enzyme binds to pyrimidine dimers
and converts them back to the original monomeric
nucleotides.

7.  Excision repair involves the excision of a segment of the polynucleotide containing a
damaged site, followed by resynthesis of the correct nucleotide sequence by a DNA
polymerase.
 These pathways fall into two categories:
1. Base –excision repair
 Base excision repair involves removal of a damaged nucleotide base, excision of a short
piece of the polynucleotide and resynthesis with a DNA polymerase.
 It is used to repair many minor damage like alkylation and deamination resulting from
exposure to mutagenic agents.
 Enzyme DNA glycosylase initiates the repair process.

8.  A DNA glycosylase does not cleave phosphodiester bonds, instead cleave the N- glycosidic
bonds, liberating the altered base and generating apurinic or an apyrimidinic site, both
called as AP sites.
 The resulting AP site is then repaired by an AP endonuclease repair pathway.
 These enzymes introduce chain breaks by cleaving the phosphodiester bonds at AP sites.
 This bond cleavage initiates an excision-repair process with the help of three enzymes_
 Exonuclease
 DNA polymerase I
 DNA ligase

10. Nucleotide excision repair
 This is similar to base-excision repair, but is not preceded by the removal of a damaged
base and can act on more substantially damaged areas of DNA.
 This repair system includes the breaking of a phosphodiester bond on either side of the
lesion, on the same strand, resulting in the excision of an oligonucleotide.
 The excision leaves a gap that is filled by repair systems, and a ligase seals the breaks.

11. Uvr system of excision repair in E. coli
 It involves the removal of relatively short, usually 12 nucleotides in length.
 The key enzyme is made up of three subunits, products of the uvrA, uvrB and uvrC
genes, and is called ABC excinuclease.
 ABC excinuclease binds to DNA at the site of a lesion.
 First uvrA and uvrB attach to the DNA at the damaged site.
 UvrA recognizes the damage.
 Departure of uvrA allows uvrC to bind, forming uvrBC dimer which cleaves the
damaged strand at the eighth phosphodiester bond on the 5’ side of the lesion and at
the fourth or fifth phosphodiester bond on the 3’ side.
 UvrD is a helicase that helps to unwind the DNA to allow release of the single strand
between the two cuts.
 The resulting gap is filled in by DNA polymerase I and sealed by ligase.

13.  The mismatch repair (MMR) system can detect mismatches that occur in DNA
replication.
 Enzyme systems involves in mismatch repair are as follows:
i) Recognize mismatched base pairs.
ii) Determine which base in the mismatch is the incorrect one.
iii) Excise the incorrect base and carry out repair synthesis.
 A mismatched base pair causes a distortion in the geometry of the double helix that can
be recognized by a repair enzyme system.
 it is important that the repair system distinguish the newly synthesized strand, which
contains the incorrect nucleotide, from the parental strand, which contains the correct
nucleotide.

14.  In E. coli, the two strands are distinguished by the presence of methylated adenosine
residues on the parental strand by Dam methylase.
 DNA methylation does not appear to be utilized by the MMR system in eukaryotes,
and the mechanism of identification of the newly synthesized strand remains unclear.
 There are three mismatch repair systems in E. coli on the basis of the relative lengths
of the unmethylated strand excised and repaired by components of the repair
system.
 These are categorized as:
 Long patch
 Short patch
 Very short patch.

15. Long patch system in E. coli
 In E. coli, long patch system involves three Mut proteins, coded by the mut genes.
 These Mut proteins are MutH, MutL, and MutS.
 Recognition of the sequence GATC and of the mismatch are specialized functions pf
the MutH and MutS proteins, respectively.
 The MutH protein cleaves the unmethylated strand on the 5’ side of the G in the GATC
sequence.
 The combined action of DNA helicase II and exonuclease I then removes a segment of
the new strand between the cleavage site and a point just beyond the mismatch.
 The resulting gap is filled in by DNA polymerase and the nick is sealed by DNA ligase.

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